Clonal hematopoiesis (CH) occurs with aging and is associated with increased mortality, a 10-fold increase of incident cancers, and a >2 fold increase in coronary artery disease (CAD) and stroke. In CH a significant proportion of circulating leukocytes are derived from a single dominant hematopoietic stem cell (HSC) lineage. Somatic mutations in oncogenic driver and other genes may be responsible. CH is also associated with shortened leukocyte telomere length (LTL) and accompanies biological aging and its effects, independent of chronological age. Healthy first degree relatives from families with early-onset CAD demonstrate premature aging cardiovascular phenotypes and bear a risk for incident CAD that is 2-12 times that of the general population. These early-onset CAD families have highly heritable shortened LTL at young ages, and higher rates of cancer. They may also have an increased prevalence of CH for their age, possibly contributing mechanistically to CAD risk. We posit that populations with increased genetic susceptibility to early CAD and shorter telomeres exhibit early-onset biological aging and have a higher prevalence of clinically silent CH, which in turn may contribute to both atherosclerosis and cancer risk. No studies have investigated this hypothesis in families with early-onset CAD, a putatively highly susceptible population. Our hypothesis is that a family history of early CAD conveys a higher prevalence of CH and shorter LTL for age, contributing mechanistically to a potent atherosclerosis substrate, with greater CAC score and higher rates of incident CAD. We will determine the prevalence of CH in 1610 subjects from GeneSTAR, healthy siblings, offspring or parents of probands with CAD < 60 years of age, using whole genome sequencing. We will determine whether CH is related to shortened LTL, to CAD events, and to CAC scores, and further whether the prevalence of CH is greater by age in subjects from families with and without early-onset CAD history in population studies (Framingham Heart Study, Multiethnic Study of Atherosclerosis, Jackson Heart Study, and the Atherosclerosis in Communities Study) in the NHLBI Trans-Omics for Precision Medicine (TOPMed) Program, of which GeneSTAR is a member. We will identify germline and somatic variants associated with LTL and determine the extent to which these are a function of family history of early CAD. Variants in candidate genes known to regulate LTL, including telomerase genes and shelterin genes will be given priority in the analysis. We are also examining increments in CH mutations and the LTL attrition rate over 6-20 years in the GeneSTAR families. For the highest priority genetic variants, we will ascertain functional significance as potential drivers of CH using gain-of-function (over-expression) and loss-of-function (gene silencing) approaches to assess their effect on proliferation and clonal expansion in cellular and in vivo murine models, and effects on atherosclerosis in Ldlr knockout mice. This novel combination of genetic epidemiology and functional studies will elucidate a potentially new mechanism for premature aging and its early-onset cardiovascular disease substrate.